Aqueous solutions of glycolic, propionic, or lactic acid in substitution of acetic acid to prepare chitosan dispersions: a study based on rheological and physicochemical properties

Abstract

Chitosan (CH) is a biopolymer derived from chitin, which is the second most abundant polysaccharide in nature, after cellulose. Their functional groups -NH₂ and -OH can form intermolecular interactions with water and other molecules, enabling a variety of applications for CH. -NH₂ groups become protonated in acidic solutions, causing an increase in electrostatic repulsion between CH chains, which facilitates their dispersion in aqueous media. Aqueous solutions of acetic acid and/or acetates buffers have been used to disperse CH, but may not be adequate for technological applications, espeacially because of the strong flavor this acid confers to formulations. In this study, 0.125; 0.250; 0.500; 0.750 and 1.000 g (100 g)⁻¹ CH dispersions were prepared in acidic aqueous media (50 mmol L⁻¹), not only with acetic (AA), but also with glycolic (GA), propionic (PA), or lactic (LA), acid aiming to evaluate the effects of biopolymer concentration and type of organic acid on: electrical conductivity, pH, density and rheological characteristics of dispersions. Moreover, ζ potential values of CH chains dispersed in these acidic aqueous media were assessed. pH, density and consistency index were influenced by the biopolymer concentration, but not by the acid type. At a given biopolymer concentration, ζ potential signs (+) and values suggested that electrostatic interactions between CH chains and counter-anions occurred, regardless of the type of the organic acid. Thus, at least from a physicochemical point of view, GA, PA or LA showed to be suitable to replace AA when preparing dispersions containing from 0.125 to 1.000 g (100 g)⁻¹ CH for technological purposes, such as thickening or stabilizer in formulated food products.

Electronic supplementary material

The online version of this article (10.1007/s13197-020-04691-0) contains supplementary material, which is available to authorized users.

Introduction

Chitosan (CH) is a biodegradable, biocompatible, non-toxic, and environmentally friendly derivate of chitin [poly-β-(1 → 4)-N-acetyl-d-glucosamine], which is the second most abundant polysaccharide in nature, after cellulose (Nidheesh and Suresh 2015; Sarbon et al. 2015). In order to produce CH, at least 50% of the acetamide groups (CH₃CONH₂) from chitin must be converted to NH₂ groups during deacetylation reaction. CH have been used in a variety of applications, including purification of water, clarification of fruit juices and beers, production of edible films, formation of microcapsules, gastroretentive drug delivery systems, and reduction of lipids absorption in the human gastrointestinal tract (Bano et al. 2017; Oryan and Sahvieh 2017; Dhumal and Sarkar 2018; Gaurav et al. 2019; Wang et al. 2019; Souza et al. 2020). Such applications are possible mainly due to the amino and hydroxyl groups present in the monomer (NH₂ group at the C2 position of each deacetylated unit, and OH groups at the C6 and C3 positions), which allow CH chains to form intermolecular interactions with water and other chemical species (Islam et al. 2017). NH₂ groups can be protonated in aqueous acid media $(\text{R}-{\text{NH}}_2+{\text{H}}^{+}\rightleftharpoons \text{R}-{\text{NH}}_3^{+})$, promoting intra and intermolecular electrostatic repulsion between CH chains. Consequently, CH molecules can be more easily hydrated, and their dispersion is favored (Shukla et al. 2013).

In addition to some above mentionned applications, research has been developed in several areas to explore other potential functionalities of this biopolymer (Ouattara et al. 2000; Goñi et al. 2017; Soares et al. 2017, 2019a). For instance, studies evaluating rheological and/or textural properties of CH have highlighted its performance as a thickener, when this polysaccharide was dispersed in solutions containing acetic acid and/or acetate buffer (Rinaudo 2006; Klinkesorn 2013; Younes and Rinaudo 2015; Hamed et al. 2016; Suleria et al. 2016). However, acetic acid unpleasant taste can be a drawback for dermatological/cosmetic or food formulations. Only few studies have been performed to study physicochemical properties of CH dispersions using other organic acid solutions rather than acetic acid for technological applications. The interest in the study of CH aqueous dispersions with other organic acids is based on their common presence in food products and cosmetics, in which they have played roles as acidifiers, preservatives, antioxidants, among others. In this way, Amorim et al. (2016) evaluated the rates of dispersion and the maximal dispersibility of CH added to aqueous solutions containing 25, 50, or 100 mmol L⁻¹ citric or lactic acid, and observed that the type of acid influenced the biopolymer dispersion, at the same acid concentration. Protonation enthalpies, ζ potential, and pHmetric/conductometric titrations of systems containing CH and each one of these two organic acids indicated different intermolecular interactions patterns between CH/citrate and CH/lactate. This study was pioneer in demonstrating that counter-anions released from different organic acids in aqueous media can influence both the dispersion rates and the maximal dispersibility of CH.

Based on these findings, Soares et al. (2019a, b) further evaluated dispersions containing 0.1 g (100 mL)⁻¹ CH prepared in aqueous media with 10, 20, 30, 40, or 50 mmol L⁻¹ acetic (AA), glycolic (GA), propionic (PA), or lactic (LA). These acids are sytematic structural variants of AA and, therefore, present a molecular similarity with it. In this study, the increase of AA, GA, PA or LA concentration: (i) reduced pH and viscosity of the dispersions, as well as |ζ potential| of CH dispersed chains; (ii) increased electrical conductivity and density of the dispersions, as well as the average hydrodynamic diameter of CH particles. Moreover, similarities in intermolecular interactions between CH and GA/LA and AA/PA counter-anions were evidenced through FT-IR analyses, explaining, with molecular features, these behaviors for the macroscopic variables studied. Furthermore, results showed that counter-anions from organic acids affected the above cited physicochemical properties, but had not significant impacts on the rheological properties of the aqueous dispersions, at the same acid concentration (Soares et al. 2019b). These authors concluded that GA, PA, or LA showed may be also suitable to replace AA in CH dispersions, considering their physicochemical characteristics. However, the understanding of how variations in the molecular structure of short chain aliphatic organic acids can affect physicochemical and techno-functional properties of CH dispersions, with concentrations greater than 0.1 g (100 mL)⁻¹, is also of great relevance to assess its actual potential as a thickening agent.

Then, this paper proposed to study physicochemical and rheological properties of CH dispersions containing different biopolymer concentrations in aqueous solutions of acetic, glycolic, propionic, or lactic acid. In addition to the controlled variation in their molecular structures, the selection of these organic acids considered their real applicability in the formulation of food and cosmetics. Lactic and propionic acids are already food additives, with no use limitation other than current good manufacturing practices (FDA 2019), being considered as generally recognized as safe (GRAS). Glycolic acid is used as both preservative in formulated products and skin lighter in cosmetic formulations (pomades, body lotions, sunscreens, and others) (Villiers et al. 1997; Mckesey et al. 2019; Liu et al. 2020). Therefore, results here reported are useful to enable the exploration of CH techno-functional properties and, consequently, to make feasible its applications in different innovative formulated products (in biotechnological, cosmetic, and food industries, for instance).

Materials and methods

Materials

Chitosan (Medium Molecular Weight, Sigma-Aldrich Corporation, USA; Product ID = 448877; Batch number = #STBF8484V) from seashell skeleton was used in all experiments. Others chemicals were of analytical grade and used without any purification process: glacial acetic acid (Vetec, Brazil; purity = 99.7%), glycolic acid (Vetec, Brazil; purity ≥ 98.0%), lactic acid (Impex Quimica, Spain; purity = 85%), propionic acid (Sigma-Aldrich Corporation, USA; purity ≥ 99.5%), and sodium acetate (Merck, Germany; purity ≥ 99.8%). Deionized water was obtained from a Mili-Q system (18.2 MΩ cm⁻¹, 25 °C; Reference A + , Millipore, Italy).

Experimental design

A factorial design 6 × 4 (totaling 24 systems) was performed to understand the effect of two independent variables (CH concentration and organic acid used in aqueous media to disperse CH) upon dependent variables (electrical conductivity, pH, ζ potential, density, consistency index, and behavior index). Six CH concentrations from 0.000 to 1.000 g (100 g)⁻¹, and four organic acids (acetic, glycolic, propionic, or lactic acid) were analysed. Treatments were replicated three times, and results were presented as average ± standard deviation.

Chitosan characterization

Before use, CH was washed three times with deionized water in order to reduce water-soluble chitooligosaccharides and salt residues. The washed CH was recovered using a vacuum filtration system and qualitative paper (Cat No 1004 125, Whatman). Then, remaining solid CH was frozen, lyophilized (Terroni, LS 3000, Brazil), and stored at 7 ± 2 °C (Consul, Pratice 410, Brazil).

Degree of deacetylation (DD)

DD was estimated by applying an Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy (ATR-FTIR) approach. CH powder was analysed using a FT-IR spectrophotometer (600-IR, Varian, USA) equipped with an attenuated reflectance accessory (GladiATR, PIKE Technologies, USA). Transmittance values (%) present in the CH spectrum were converted to absorbance values and normalized relatively to the highest corrected absorbance. Then, two regions (850–1750 cm⁻¹ and 1750–3750 cm⁻¹) were created, and the absorbances of peaks present in each region were calculated through deconvolution in Lorentzian components by PeakFit (v. 4.12, SeaSolve Software Inc., 1999–2003). Degree of acetylation (DA) was estimated using the empirical relationship between normalized absorbances of the peaks at wavenumbers 1320 and 1420 cm⁻¹, as presented in Eq. 1 (Brugnerotto et al. 2001; Kasaai 2008). Finally, DD was obtained by simple difference [DD(%) = 100% − DA(%)].

$$\text{DA}\left(\%\right)=\frac{\left(\frac{A_{1320}}{A_{1420}}-0.3822\right)}{0.03133}$$ 1

In Eq. (1), A₁₃₂₀ and A₁₄₂₀ are the normalized absorbances of the peaks at wavenumbers 1320 cm⁻¹ and 1420 cm⁻¹, respectively.

Viscometric average molar mass ${\overline{M}}_V$.

CH molar mass was estimated by viscometric-average molar mass ${\overline{M}}_V$ approach. 0.02, 0.04, 0.06, 0.08, and 0.10 g (100 g)⁻¹ CH was dispersed in acetic acid-sodium acetate buffer (0.2 mol L⁻¹ acetic acid and 0.1 mol L⁻¹ sodium acetate; pH = 4.41 and ionic strength = 0.1 mol L⁻¹) (Kasaai 2007). CH dispersions were placed in a Cannon–Fenske viscometer (model 513 20, Schott, Germany), and their flow times were measured. From these experimental data, average intrinsic viscosity ($\left[\overline{\eta }\right]$) as calculated as the average between the Huggins ([η]_H) and Kraemer ([η]_K) intrinsic viscosities (Nieto Galván et al. 2018), which were obtained by extrapolating Eqs. 2 and 3, respectively, to infinite dilution ([CH] → 0).

$$\frac{{\eta }_{\mathit{sp}}}{c}={[\eta ]}_H+k_1{[\eta ]}^{2}c$$ 2

$$\frac{ln({\eta }_r)}{c}={[\eta ]}_K+k_1^{\prime }{[\eta ]}^{2}c$$ 3

In Eqs. 2 and 3, k₁ is the Huggins constant, k’₁ is the Kraemer constant, and c is the concentration of chitosan in the diluted dispersions.

Then, viscometric-average molar mass ${\overline{M}}_V$ was calculated by the Mark–Houwink–Sakurada (MHS) relationship (Kasaai 2007; Costa et al. 2017).

$$\left[\overline{\eta }\right]=K_{\mathit{MHS}}·{\overline{M}}_V^a$$ 4

In Eq. (4), a and K_MHS are the constants of MHS relationship.

Preparation of acidic aqueous dispersions of chitosan

CH stock-dispersions (CSD) containing 1.000 g (100 g)⁻¹ CH were produced adding up appropriate amounts of this biopolymer to acid solutions (50 mmol L⁻¹), which were previously prepared using acetic (AA), glycolic (GA), propionic (PA), or lactic (LA) acid. The proposed acid concentrations aim to achieve the common acidity found in food and cosmetic products. The resulting systems were stirred during 4 h within a thermostatic bath (TE-184, Tecnal, Brazil) at 25.0 ± 0.1 oC, using a mechanical agitator (MA-039, Marconi, Brazil) equipped with a helical propeller (270 rpm). Then, CSD were centrifuged (7830 rpm; 5430, Eppendorf, Germany) during 20 min in order to remove any non-dispersed particles. CH diluted-dispersions (CDD) were prepared by CDS dilution to 0.125, 0.250, 0.500, and 0.750 g (100 g)⁻¹ using the same solution (50 mmol L⁻¹) for each acid as a diluent. Additionally, control systems (CS) without chitosan (0.000 g·(100 g)⁻¹) were prepared to each one of the organic acids. CSD, CDD and CS were transferred to amber bottles and stored at 7 ± 2 °C.

Evaluation of physicochemical properties

Electrical conductivity, pH, and density

0.000, 0.125, 0.250, 0.500, 0.750, and 1.000 g (100 g)⁻¹ CH dispersions had their electrical conductivity (Termo, Orion 145A + , USA; 25.0 ± 1.0 °C), pH (digital potentiometer Hanna, H2221, USA; 25.0 ± 1.0 °C), and density (oscillatory densimeter Schmidt Haensch, EDM, Germany; 25.00 ± 0.05 °C) measured, soon after their preparation.

ζ potential

ζ potential of dispersed CH chains was calculated from the electrophoretic mobility (Zetasizer Nano-ZS, Malvern Instruments, United Kingdom), at 25.0 ± 0.1 °C. Dispersions containing from 0.125 to 1.000 g (100 g)⁻¹ CH were placed in a folded capillary cell (DTS1070; Malvern Instruments, United Kingdom), which is required for ζ potential analyses by Zetasizer Nano-ZS. Then, the folded capillary cell was subjected to a controlled electric field, and electrophoretic mobility was estimated from the speed and the direction of the particle movement due to this electric field (Eq. 5).

$${\mu }_e=\frac{v}{\left|\overrightarrow{E}\right|}$$ 5

In Eq. 5, μ_e is the electrophoretic mobility, v is the speed of dispersed CH chains, and $\overrightarrow{\text{E}}$ is the electric field.

ζ potential values were calculated considering the Smoluchowski model for the double electrical layer (Eq. 6).

$${\mu }_e=\frac{{\epsilon }_r{\epsilon }_o\zeta }{\mu }$$ 6

In Eq. 6, ε_r is the dielectric constant of the medium, ε_o is the permittivity of free space, ζ is the zeta potential, and μ is the viscosity of the acid aqueous dispersion.

Rheological properties

Rheological measurements of CH dispersions, containing from 0.125 to 1.000 g (100 g)⁻¹ and prepared with acetic, glycolic, propionic, or lactic acid solutions, were performed using a rotational rheometer (Discovery Hybrid Rheometer 1, TA Instruments, USA), equipped with a stainless steel parallel plate sensor (diameter = 25 mm; gap = 1 mm), at 25.0 ± 0.1 °C. Flow curves were determined by progressively varying the shear rate $\dot{\gamma }$ from 0.1 to 200 s⁻¹ in three cycles (1st up cycle, down cycle, and 2nd up cycle; 300 s each cycle), and measuring the corresponding shear stresses (τ).

Statistical analyses

Statistical analyses were performed using the SAS software (version 9.3, SAS Institute Incorporation, USA). Power law model (Eq. 7) was fitted to $\tau =f(\dot{\gamma })$ experimental data. Coefficient of determination (R²) and mean absolute percentage error (MAPE) were used to evaluate the adequacy of fitting in all cases.

$$\tau =K·{\left(\dot{\gamma }\right)}^n$$ 7

In Eq. 7, K is the consistency index, and n is the flow behavior index.

Dependent variables (electrical conductivity, pH, density, consistency index, and behavior index) of aqueous media containing different organic acids along with ζ potential values of dispersed CH chains were compared by ANOVA (p < 0.05), at a given biopolymer concentration. When pertinent, Dunnett’s test (p < 0.05) was utilized to compare CH dispersions containing acetic acid with others.

Results and discussion

Chitosan characterization

Some variability is expected in degree of polymerization, molar mass, and hydrodynamic diameter distribution of polysaccharides (Dash et al. 2011; Knidri et al. 2018). In the specific case of CH, variations in the biological origin, conditions used in the deacetylation reaction, and degree of acetylation have been causing modifications in dispersion kinetics, maximum dispersibility, and techno-functional properties (Hamed et al. 2016; Knidri et al. 2018). Thus, a previous characterization of CH has been recommended in order to discuss modifications on physicochemical and rheological aspects of dispersions as a function of the CH molecular characteristics.

In Fig. 1a, ATR-FTIR spectra of CH between 850 and 1750 cm⁻¹ was used to estimate the CH degree of deacetylation (DD). Upon replacing intensities values of peaks at wavenumbers 1320 cm⁻¹ and 1420 cm⁻¹ in the Eq. 1, DD value of the CH was estimated as 72.5% (Brugnerotto et al. 2001; Kasaai 2008). During the deacetylation reaction to obtain CH, chitin is exposed to strongly alkaline reaction media at high temperatures (~ 150 °C) and, as a result, chitooligosaccharides (CH with degrees of polymerization ≤ 20 and DD usually ≥ 95.0%) can also be produced (Hosseinnejad and Jafari 2016). Chitooligosaccharides should be eliminated during the CH washing, causing a reduction of DD% informed by the manufacturer (83.2%) (2019). DD value influences CH dispersibility in acid aqueous media, wherein DD values ≥ 70.0% have been correlated to high dispersion rates (Younes and Rinaudo 2015; Islam et al. 2017). In this case, checking experimentally the DD of commercial CH prior to usage can be a practical strategy to know its characteristics and predict its techno-functional applications.

Fig. 1.

FT-IR spectra from chitosan to 850–1750 cm⁻¹ (a): absorbance from FT-IR spectra (∙∙) and absorbance calculated through deconvolution in Lorentzian components (-); and extrapolation to infinite dilution [(chitosan) → 0] of Huggins and Kraemer empirical models adjusted to viscometric-average experimental data from chitosan aqueous dispersions b: (●) $\frac{{\eta }_{\mathit{sp}}}{c}=k_1{[16.3]}^2·c+[6.3];R^2=0.999$, (○)$\frac{ln({\eta }_r)}{c}=k_1^{\prime }{[-4.9]}^2·c+[6.4];R^2=0.991$ (b)

The extrapolation to infinite dilution ([CH] → 0) of Huggins and Kraemer empirical models used to calculate ${\overline{M}}_V$ was presented in Fig. 1b. Intrinsic viscosity of CH dispersed in acetic acid-sodium acetate buffer was estimated as $[\overline{\eta }]$ = 6.4 dL g⁻¹, from the simple average between [η]_H = 6.3 dL g⁻¹ and [η]_K = 6.6 dL g⁻¹. After DD % estimation, the constants a = 0.93 and K_MHS = 3.6.10^–5 dL g⁻¹ of MHS relationship (Eq. 4) could be obtained (Kasaai 2007). Then, viscometric-average molar mass ${\overline{M}}_V$ of CH was calculated as 430 ± 30 kDa, which is in accordance with the specification to molecular weight of CH presented by Sigma-Aldrich. The increase in molar mass has been correlated to the raise in dispersion viscosities, (Klinkesorn 2013), and, for CH products with molar mass > 300 kDa, is expected a good performance as a thickener and stabilizer (Laplante et al. 2005; Klinkesorn 2013; Sigma-Aldrich 2019). Furthermore, CH dispersions viscosities are strongly dependent on the acid solutions/buffers used, which can influence biopolymer-biopolymer and/or biopolymer-water interactions in the colloidal network (Calero et al. 2010, 2013; Klinkesorn 2013).

Physicochemical properties

1.000 g (100 g)⁻¹ CH added to solution of acetic, glycolic, propionic, or lactic acid (50 mmol L⁻¹) was fully dispersed when visually assessed. Then, CSD were diluted with the same acid solution used in the preparations (50 mmol L⁻¹) in order to obtain CDD. Electrical conductivity, pH and density of CH dispersions were analysed and results were presented in Fig. 2 along with ζ potential values of dispersed CH chains. At a given biopolymer concentration, no significant differences were observed by ANOVA (p > 0.05) in the above mentioned variables.

Fig. 2.

Electrical conductivity (a), pH (b), and density (d) of aqueous media containing different chitosan concentrations dispersed in acetic ( ), glycolic ( ), propionic ( ) or lactic ( ) acid solutions; ζ potential of chitosan dispersed particles (c) present in 0.125 g (100 g)⁻¹, 0.250 g (100 g)⁻¹, 0.500 g (100 g)⁻¹, 0.750 g (100 g)⁻¹, and 1.000 g (100 g)⁻¹ chitosan dispersion containing acetic ( ), glycolic ( ), propionic ( ), or lactic ( ) acid. *At a given CH concentration, no significant differences were observed to aqueous dispersions by ANOVA (p > 0.05)

pH values for CH dispersion containing different organic acid were showed in Fig. 2b. pH values of CH dispersions increased (from 3.5 ± 0.2 to 5.1 ± 0.1 for systems containing AA; 2.9 ± 0.1 to 4.3 ± 0.1 for GA; 3.8 ± 0.1 to 5.1 ± 0.1 for PA; and 2.8 ± 0.1 to 4.8 ± 0.1 for LA) as biopolymer concentration augmented from 0.125 to 1.000 g (100 g)⁻¹. This observation can be explained based on the protonation of NH₂ groups by H⁺ cations present in aqueous media, i.e., as the amount of chitosan increased, there were more NH₂ to be protonated and, therefore, less free H⁺ in the bulk (Shukla et al. 2013). As expected, control systems (CS) presented pH values (AA = 3.1 ± 0.1; GA = 2.6 ± 0.1; PA = 3.2 ± 0.1; and LA = 2.5 ± 0.1) lower than CH dispersions. At a given CH concentration, dispersions containing AA or PA presented higher pH values, comparing to what was observed for those prepared with GA or LA. The inductive effect acting on carboxylic acids molecular structure could explain this observation: when compared to AA or PA molecular structures, the carbon atom of the carboxyl group present in GA or LA supports a higher positive charge due to the hydroxyl group attached to their C2 carbon; thus, OH group exerts an attraction on electronic clouds of the molecule towards the oxygen atom of this hydroxyl group, and protons from GA or LA are more easily released by COOH groups (Siggel and Thomas 1988; Siggel et al. 1986). As a consequence of such structural molecular characteristics, the pK_a values of these organic acids (AA = 4.75; GA = 3.83; PA = 4.87; and LA = 3.85) is affected, which in turn, facilitates the release of H⁺ (Solomons and Fryhle 2000).

Electrical conductivity for CS and CH dispersions were presented in Fig. 2a. Both H⁺ cations and the counter-anions are excellent electrical conductors, and electrical conductivity values for CS (AA = 435 ± 5 μS cm⁻¹; GA = 1254 ± 38 μS cm⁻¹; PA = 359 ± 19 μS cm⁻¹; and LA = 1302 ± 16 μS cm⁻¹) are related to the number of deprotonated organic acids molecules. As expected, aqueous media containing GA or LA showed a higher density of electrically charged chemical species. Electrical conductivity values increased in aqueous media containing AA (from 443 ± 5 to 1103 ± 34 μS cm⁻¹) or PA (from 362 ± 23 to 965 ± 23 μS cm⁻¹) as the CH concentration was raised. On the other hand, a lower variation was observed for GA (from 1010 ± 46 to 1381 ± 50 μS cm⁻¹) or LA (from 953 ± 50 to 922 ± 11 μS cm⁻¹), in the same conditions. These observations indicate that CH added to aqueous media containing AA or PA causes a more pronounced equilibrium shift towards the acid molecules deprotonation. As stated previously, an expressive increase in pH values was observed as CH concentration was raised in aqueous media containing both AA and PA, which pointed out that a great amount of protons was bound to NH₂ groups (there were less H⁺ free in the bulk). Thus, the AA and PA were deprotonated in order to reestablish a new chemical equilibrium to the system. On the other hand, GA or LA presented a more intense deprotonation in aqueous media without CH, promptly providing a higher amount of H⁺ to NH₂ groups to biopolymer chains protonation. Then, a lower variation in electrical conductivity values observed for aqueous media containing GA or LA, which suggested a lower deprotonation of these acids as CH concentration was increased. Similar to what was said for the pH results, the observations for electrical conductivity were in agreement with the explanation about the inductive effect acting on carboxylic acids molecular structure.

ζ potential of dispersed CH chains were also measured, and results were represented in Fig. 2c. CH chains dispersed in all four organic acids showed similar ζ potential values (from + (70.0 ± 5.0) to + (81.2 ± 2.0) mV for systems containing AA; + (71.7 ± 2.0) to + (80.8 ± 5.0) mV for GA; + (72.6 to ± 8.0) to + (78.7 ± 4.0) mV for PA; and + (66.5 ± 2.0) to + (80.4 ± 2.0) for LA), and as expected, they were all positive. In addition, the type of acid added to aqueous medium did not alter dramatically ζ potential values of CH chains, at the same biopolymer concentration. These observations indicated that similar interactions between CH chains and the organic acid counter-anions were established. Soares et al. 2019b observed that the increase in the counter-anions concentration around the CH chains attenuated the density of positive charges on the electrical layer of the dispersed biopolymer chains, leading to smaller ζ potential values. Furthermore, at a constant organic acid concentration, ζ potential values showed that the protonation of NH₂ group was not influenced by the distinct counter-anions.

According to Fig. 2d, CS showed similar values for density (AA = 0.9971 ± 0.0004 g cm⁻³; GA = 0.9980 ± 0.0006 g cm⁻³; PA = 0.9969 ± 0.0005 g cm⁻³; and LA = 0. 9973 ± 0.0002 g cm⁻³). In addition, CH dispersions presented an increase for these values (0.9978 ± 0.0005 to 1.0013 ± 0.0001 g cm⁻³ in systems containing AA; 0.9987 ± 0.0002 to 1.0022 ± 0.0004 g cm⁻³ for GA; 0.9974 ± 0.0003 to 1.0011 ± 0.0003 g cm⁻³ for PA; and 0.9980 ± 0.0008 to 1.0013 ± 0.0005 g cm⁻³ for LA), as the biopolymer concentration was augmented from 0.125 to 1.000 g (100 g)⁻¹. Noticeably, the increase of the dispersion density values was analogous for all acids, which corroborates the assumption that the interaction between the counter-anions and the CH chains is quite similar.

Briefly, CH added to acid aqueous media altered the equilibrium of protonated-deprotonated chemical species, according to electrical conductivity and pH measurements. Furthermore, this effect was more pronounced in systems containing AA or PA. At a given biopolymer concentration, alterations in the acid deprotonation equilibrium did not influence expressively the protonation of CH chains (according ζ potential values) nor the balance of interactions chitosan-water and chitosan-chitosan (according density values). Modifications in the colloidal network and dispersions physical properties have been reported for systems wherein alterations in protonation or electrostatic shielding could be observed (Rinaudo 2006).

Rheological properties

Rheograms of CH dispersions were analysed, and hysteresis was not observed (curves corresponding to up-down-up cycle were superposed). When visually assessed, the flow curves showed a pseudoplastic behavior, and τ₀ next to 0 Pa (rheograms of 2nd up curves were presented in Figure SM1). Then, $\tau =f(\dot{\gamma })$ data from 2nd up curve was used to fit the Power Law model. Power Law model was well-fitted to experimental data, presenting R² values ≥ 0.95 and MAPE values ≤ 5% (adequacy fitting for the Power Law model adjusted to experimental data were showed in Table SM3). Consistency index and behavior index of CH dispersions obtained by the Power Law model were presented in Fig. 3. At a given biopolymer concentration, no significant differences were observed to CH dispersions containing different organic acids by ANOVA (p > 0.05), in terms of consistency index and behavior index.

Fig. 3.

Consistency index and behavior index for the Power Law model adjusted to chitosan dispersions prepared in aqueous media containing acetic ( ), glycolic ( ), propionic ( ), or lactic ( ) acid. *At a given CH concentration, no significant differences were observed to aqueous dispersions by ANOVA (p > 0.05)

Consistency values exponentially increased according CH concentrations was augmented from 0.125 to 1.000 g (100 g)⁻¹ (0.11 ± 0.01 to 1.19 ± 0.03 Pa·sⁿ for systems containing AA; 0.12 ± 0.01 to 1.13 ± 0.04 Pa·sⁿ for GA; 0.11 ± 0.01 to 1.11 ± 0.04 Pa·sⁿ for PA; and 0.12 ± 0.01 to 1.08 ± 0.04 Pa·sⁿ for LA) in acid aqueous media. The augment in CH concentration induced modifications in physical properties of aqueous media, which could be observed by the exponential increase in consistency index values. Moreover, small differences were seen for CH dispersions prepared with the different acids, which indicated that intermolecular interactions between water molecules and chains CH were not affected by the type of counter anion present.

Behavior index values increased as CH concentration was raised from 0.125 to 0.500 g (100 g)⁻¹ in aqueous media (0.57 ± 0.02 to 0.72 ± 0.03 for systems containing AA; 0.53 ± 0.02 to 0.72 ± 0.03 for GA; 0.58 ± 0.02 to 0.73 ± 0.03 for PA; and 0.52 ± 0.02 to 0.72 ± 0.04 for LA). The alignment of biopolymer chains as the shear rate increased has been reported to augment behavior index values, which could promote the existence of a behavior closer to Newtonian flow under critical conditions (Fischer et al. 2009; Rao 2013). This hypothesis can be supported by the moderate rigidity, lack of branch, and medium/high molar mass characteristics of the CH chains (Islam et al. 2017; Kaisler et al. 2020). In this case, higher values of n could indicate the restructuration of dispersed particles at a molecular level, representing a response to higher shear rate values, and causing the decrease of chitosan-chitosan interactions (by their alignment) and consequent an increase in water-biopolymer molecular interactions. On the other hand, an increase for behavior index values in CH dispersions containing higher biopolymer concentrations (0.70 ± 0.01 and 0.64 ± 0.01 for aqueous media prepared respectively with 0.750 and 1.000 g (100 g)⁻¹ CH containing AA; 0.69 ± 0.01 and 0.65 ± 0.01 for GA; 0.71 ± 0.01 and 0.65 ± 0.01 for PA; and 0.70 ± 0.01 for 0.65 ± 0.01 for LA) was not notated. These observations could be explained by the formation of chitosan chains microscopic aggregates or by the growth of the particles within acid aqueous solution as the shear rate was increased in CH dispersions containing higher biopolymer (0.750 to 1.000 g (100 g)⁻¹), contributing to increase of the pseudoplasticity (Martínez et al. 2004; Calero et al. 2010). Moreover, the highest values for consistency index were also noticed in these systems, which could also indicate the partial aggregation of CH chains. Furthermore, the type of organic acid used to prepare CH dispersions seems not to have influenced the behavior index values, at a given biopolymer concentration.

Few studies have reported the use of different acids to produce chitosan dispersions in order to prepare materials (such as polymeric films and edible coats for fruits and vegetables), and to achieve some specific functions in the products (antimicrobial, antioxidant, or acidifying agent, for example). However, these studies did not highlight the effects of replacing or combining acetic acid with other organic acids on the formation of aqueous dispersions and/or on the physicochemical characteristics of the dispersions obtained. Furthermore, they did not evaluate the consequent impacts on the structure and properties of coated /packed foods (Ouattara et al. 2000; Park et al. 2002; Kim et al. 2006; Badawy and Rabea 2009; Romanazzi et al. 2009; Campos et al. 2011; Ferreira et al. 2020). Indeed, it is important to understand the influence of the type of acid used on physicochemical and rheological properties of chitosan dispersions containing different biopolymer concentrations, before designing any food application to them. Then, our research aimed to evaluate the effects of replacing the most often used acetic acid by other food grade organic acids. Since no expressively variations were observed for the analyzed macroscopic physicochemical characteristics (electrical conductivity, pH, density and rheological parameters of dispersions, along with ζ potential values of dispersed chitosan chains), results here reported indicate that GA, PA, or LA showed to be suitable to replace AA when preparing aqueous dispersions containing from 0.125 to 1.000 g (100 g)⁻¹ CH. Therefore, our present study was planned to clarify such physicochemical and rheological properties of chitosan dispersions, widening the possibilities of replacing acetic acid by other food-grade organic acids.

Conclusion

Chitosan (CH) dispersions were successfully prepared in aqueous media containing AA, GA, PA, or LA. Aqueous media containing AA or PA acid presented more intense alterations for electrical conductivity and pH values as the CH concentration was raised, indicating a higher alteration of the equilibrium of protonated-deprotonated chemical species than in systems prepared with GA or LA. However, density, consistency and behavior indexes were not influenced by these alterations in the equilibrium of protonated-deprotonated acid molecules, at a given biopolymer concentration. The same can be said about ζ potential values of chitosan chains dispersed in acid aqueous media. Then, the mentioned variables were influenced by the CH but did not depend on the type of acid. In other words, the type of acid seems not to have caused any mischief to CH dispersibility, nor to the balance of interactions chitosan-water and chitosan-chitosan, at a given biopolymer concentration. Results presented in this study bring new information about CH dispersions prepared in aqueous media with other technologically relevant organic acids rather than AA. In summary, GA, PA, or LA showed to be suitable to replace AA when preparing dispersions containing higher concentrations of CH (up to 1.000 g (100 g)⁻¹), widening the applicability of this biopolymer in food or cosmetic formulations, as an alternative thickening/stabilizing agent.

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Abbreviations and symbols

a

First constant of Mark–Houwink–Sakurada relationship (dimensionless)

AA

Acetic acid

A₁₃₂₀

Normalized absorbance of the peak at wavenumbers 1320 cm⁻¹ (norm. arb. u.)

A₁₄₂₀

Normalized absorbance of the peak at wavenumbers 1420 cm⁻¹ (norm. arb. u.)

CDD

Chitosan diluted-dispersions

CH

Chitosan

CS

Control systems

CSD

Chitosan stock-dispersions

DA

Degree of acetylation (%)

DD

Degree of deacetylation (%)

GA

Glycolic acid

K

Consistency index (Pa sⁿ)

K_MHS

Second constant of Mark–Houwink–Sakurada relationship (dL g⁻¹)

LA

Lactic acid

MAPE

Mean absolute percentage error (%)

${\overline{M}}_V$

Viscometric-average molar mass (kDa)

n

Flow behavior index (dimensionless)

PA

Propionic acid

pH

Hydrogenionic potential (dimensionless)

R²

Coefficient of determination

ζ

Zeta potential (mV)

μ

Viscosity (Pa s)

μ_e

Electrophoretic mobility (m² (V s)⁻¹)

ν

Speed of particles (m s⁻¹)

τ

Shear stress (Pa)

$\dot{\gamma }$

Shear rate (s⁻¹)

$\overrightarrow{\text{E}}$

Electric field (N C⁻¹)

[η]_H

Huggins intrinsic viscosity (dL g⁻¹)

[η]_K

Kraemer intrinsic viscosity (dL g⁻¹)

$\left[\overline{\eta }\right]$

Average intrinsic viscosity (dL g⁻¹

Author contributions

Lucas de Souza SOARES: Conceptualization, Methodology, Investigation, Formal Analysis, Validation, Writing - Original Draft, Writing - Review & Editing. Bruna TONOLE: Conceptualization, Investigation, Writing - Original Draft. Gustavo Leite MILIÃO: Conceptualization, Investigation, Writing - Original Draft. Alvaro Vianna Novaes de Carvalho TEIXEIRA: Conceptualization, Writing - Review & Editing. Jane Sélia dos Reis COIMBRA: Conceptualization, Writing - Review & Editing, Resources, Funding acquisition. Eduardo Basílio de OLIVEIRA: Conceptualization, Validation, Writing - Original Draft, Writing - Review & Editing, Supervision, Project administration, Funding acquisition.

Compliance with ethical standards

Conflict of interest

In compliance with Ethical Requirements of the Journal of Food Science and Technology, we inform that all authors are aware with the submission of the present manuscript to JFST. Thus, the authors declare that there is no conflict of interest regarding the publication of this manuscript.